Programmed death 1 (PD-1) is a member of the CD28 family of receptors comprising CD28, CTLA-4, PD-1, ICOS, and BTLA (Freeman et al. (2000) J Exp Med 192:1027-34; Latchman et al. (2001) Nat Immunol 2:261-8). PD-1 is an inducible immunosuppressive receptor mainly upregulated on activated T cells and B cells during the progression of immunopathological conditions. PD-1 interaction with its ligand PD-L1 results in the inhibition of TCR and BCR mediated proliferation and cytokine production and induction of apoptosis of antigen specific T cells through the intrinsic PD-1 mediated negative signaling of an immunoreceptor tyrosine-based inhibitory motif (ITIM) (Agata et al. (1996) Int. Immunol. 8:765, Unkeless and Jin. (1997) Curr. Opin. Immunol. 9:338-343, Okzaki et al. (2001) PNAS 98:13866-71, Dong et al. (2002) Nat. Med. 8:793-800). PD-L1 is a cell surface glycoprotein and a major ligand for PD-1. PD-L1 is also inducible on lymphoid tissues and non-lymphoid peripheral tissues following cellular activation. PD-L1 is upregulated in a variety of affected cell types including cancer and stromal cells in addition to immune cells, and plays an active role in immunosuppression during the course of the deterioration of diseases (Iwai et al (2002) PNAS 99:12293-7, Ohigashi et al. (2005) Clin Cancer Res 11:2947-53). PD-L1 upregulation has been linked to poor clinical outcomes in a variety of cancers and viral infection (Hofmeyer et al. (2011) J. BioMed. Biotech. 2011:1-9, McDermott and Atkins. (2013) Cancer Med. 2:662-73). The blockade of PD-1 or PD-L1 by antibody promoted CD8 T cell infiltration, CTL activity and increased presence of Th1 cytokine IFN-gamma in preclinical and clinical settings (Zhou et al. (2010) J. Immunol. 185:5082-92, Nomi et al. (2007) Clin Cancer Res. 13:2152-7, Flies et al. (2011) Yale J. Bio. Med. 48:409-21, Zitvogel and Kroemer. (2012) Oncolmmunol. 1:1223-25). PD-L1 antibody as an immunomodulating agent has been shown to be efficacious when used as monotherapy or combined with antibodies to other immunosuppressive molecules.
However, the immunomodulating intervention to immunosuppressive factors only partially resolves the problems associated with impaired immunity in cancer, infection, and other diseases. It is still highly desirable to utilize biotherapeutic agents to directly stimulate and expand effector immune cells for lifting weakened innate and adaptive immune response to a more effective level to control tumor and infection. Immunotherapy using cytokines including interleukins, i.e. IL-2, IL-12, IL15, IL-21, and TNFα, GM-CSF, etc., has been shown to be efficacious to some extent in the treatment of cancer and infection, but clinical outcome is often limited by systemic toxicity associated with the high blood concentrations of cytokine that need to obtain efficacy and lack of specificity of target in affected cells and tissues.
Among assessed cytokines, IL15 has been recognized to be dedicated to stimulate effector and central memory CD8 T cells composing of a subset of antigen specific CD8 cells to exert antitumor immunity without modulating effects on other T cell populations. Moreover, unlike IL-2 that activates Treg, IL15 has been shown to have capacity to rescue T cells from apoptosis induced by Treg and other immunosuppressive cells in addition to its ability to activate natural killer (NK) cells and effector and memory CD8 T cells (Van Belle et al. (2012) PLoS One 7:e45299, Obar and Lefrancois. (2010) J. Immunol. 185:263-72, Pelletier and Girard. (2006) J Immunol 177:100-108, Elpek et al. (2010) PNAS 107:21647-21652).
IL15 was identified as a yc cytokine in 1994 based on its ability to stimulate the proliferation of the murine T cell line CTLL-2 (Grabstein et al. (1994) Science 264:965-8, Bamford et al. (1996) PNAS 93:2897-902). Human IL15 shares approximately 97% and 96% amino acid sequence identity with simian and cynomolgus IL15, respectively. Human and mouse IL15 have 73% homology and are comparably active on mouse cells. IL15 is a 12.5 KD protein (114 amino acids), secreted by DC, macrophage and granular cells as a 14-15 kDa glycoprotein, and also a member of the four α-helix bundle-containing cytokines (Andderson et al. (1995) J Biol Chem. 270:29862-9, Steel et al. (2012) Trends Pharmacol. Sci. 33:35-41). IL15 is typically formed a complex with IL15 receptor alpha expressed on APCs prior to binding to functional IL15 receptor beta and gamma units on T cells and NK cells. IL15 may be presented in trans to responsive cells expressing CD122 and CD132 by cells expressing the cytokine itself bound to a membrane form of the receptor alpha chain (Dubois et al. (2002) Immunity 17:537-47). IL15 receptor alpha sushi domain (29.5 KD in size) is a critical component to form a complex with IL15 prior to properly engagement with receptor β and γ (Wei et al. (2001) J. Immunol. 167:277-82). IL15 and IL15Rα complex and IL15/IL15Rα sushi domain fusion protein were reported to be highly potent to stimulate CD8 T cells and NK cells in vitro and in vivo compared to IL15 alone (Mortier et al. (2005) J Biol Chem. 281:1612-19, Stoklasek et al. (2006) J. Immunol. 177:6072-80). IL15 also induces the proliferation and differentiation of stimulated human B cells (Armitage et al. (1995) J Immunol. 154:483-90). It was suggested that IL15 mostly opposed activation-induced cell death (AICD) by acting to prolong the survival of T lymphocytes (Marks-Konczalik et al. (2000) PNAS 97:11445-50). IL15 has an exceptional ability to support the maintenance of NK cells and memory phenotype and antigen specific memory CD8 T cells (Ma et al. (2006) Annu Rev Immunol. 24:657-79). Thus, among most active cytokines in immunomodulation, IL15 has an unique capacity to mediate many important aspects of immunity against a variety of tumor types and viral infection including HIV, HBV, HCV, LCMV, etc (Steel et al. (2012) Trends Pharmacol. Sci. 33:35-41, Verbist and Klonowski, (2012) Cytokine. 59:467-478).